Methods:
ASC oxidation by UVA photolysis (100 mW/cm2 for 2 hrs) of kynurenines or enzyme digested water insoluble proteins was assessed by spectrophotometry. The UV irradiated and dark control samples were incubated with human lens proteins at pH 7.4 and 37°C to determine formation of AGEs. Pentosidine, a representative marker for AGEs, was quantified by reverse phase HPLC.

Results:
UVA photolysis of mixtures containing kynurenine (N-formyl kynurenine-NFK or kynurenine-Kyn or 3OH-Kynurenine-3OHKyn) and ASC showed that NFK aggressively promoted oxidation of ASC. ASC oxidation occurred from both free and protein bound kynurenines (in the presence or absence of oxygen). Addition of glutathione (4 mM) or lens proteins (2 mg/ml) did not prevent such kynurenine-mediated ASC oxidation. Oxidation of ASC was also apparent in the UVA irradiated water-insoluble protein. Cataractous lens proteins digest induced ASC oxidation at a significantly higher rate than the age matched normal lens proteins. Incubation of UVA-irradiated kynurenine-modified lens proteins or UV-irradiated mixtures of ASC and kynurenines for 7 days at 37°C resulted in the formation of pentosidine, an AGE present in human lens protein. These data show that UVA photolysis of kynurenines (free and protein-bound) and lens pigments bring about oxidation of ASC, which results in AGE formation in lens proteins.

Conclusions:
Kynurenines promote ASC oxidation and such oxidation is not dependent on molecular oxygen. Kynurenine-mediated ASC oxidation and subsequent AGE formation might play an important role in lens protein crosslinking and aggregation during aging and cataract formation in humans.